Identification And Exploration Of Type 1 Diabetes Susceptibility Genes And Functional Mechanisms

Type 1 diabetes mellitus(T1DM) is one of the major threats for public health. It is a serious disorder characterized by destruction of pancreatic β cell, cumulating to absolute insulin deficiency. Genetic factors play an important role in the development of T1 DM. Genome-wide association studies(GWAS) have identified many susceptible genes for T1 DM, but more with minor effect remained unknown, and could only be uncovered via more powerful analytical approach. Additionally, most of the current availlable GWAS have just established statistical association between disease and genetic variation at DNA level, while the biological and functional mechanism for the association is largely unknown. Therefore, the aim of the present study is to identify additional susceptible T1 DM genes through a powerful approach, and to explore functional mechanisms of the identified associations. Specifically, this study is composed of two parts: Part 1, performing powerful Gene-based GWAS, replication study and function prediction analysis to identify novel susceptibility genes for T1DM; Part 2, conducting integrative study of integrated gene relationships among implicated loci(GRAIL) analysis, differential gene expression analysis, functional prediction and annotation clustering analysis and combined with expression quantitative trait loci(e QTLs) result to further explore functional mechanisms of the identified genes.Part 1Objective: To identify novel susceptibility genes for T1 DM through powerful Gene-based GWAS.Materials and Methods: This study conducted the discovery Gene-based GWAS in a total of 4,075 T1 DM cases and 2,604 controls with GATES algorithm implemented in KGG, and replicated significant genes in the different populations. This study further conducted differential expression analysis and functional annotation clustering analysis to validate the identified genes.Results: This study have identified 452 genes associated with T1 DM at the Gene-based genome-wide significance level(P=9.05E-04). Two-hundred and eighty one of the identified genes have been previously reported according to previous study, corroborating the validity of our analytical approach. Of the remaining 171 novel genes, 53 genes were successfully replicated in different populations and/or differential expression studies. Therein, 7 genes were replicated in different population and had differential expression in peripheral blood monocyte cells(PBMC) or monocyte. Detailed annotation of these 7 genes showed that 3 genes locate in the HLA regions and the remaining 4 genes(RASIP1, STRN4, BCAR1 and MYL2) locate outside the HLA region. Functional clustering analysis showed that all the identified genes enrich in immune related pathways or GO terms.Conclusion: By using powerful Gene-based GWAS and replication analysis, this study have identified 53 novel susceptible genes for T1 DM with supportive evidence form replication studies and/or differential expression studies, establishing solid basis for uncovering genetic and functional mechanism for the disease.Part 2Objective: To explore functional mechanisms of T1 DM associtated genetic variants through integrative analyses of multiple-omics data and functional prediction analysis.Materials and Methods: Based on publicly available datasets, we conducted integrative analyses(i.e., integrated gene relationships among implicated loci(GRAIL), differential expression analysis, functional prediction and functional annotation clustering analysis) and combined with expression quantitative trait loci(e QTL) result to further explore functional mechanisms underlying the development of T1 DM.Results: This study identified 17 T1DM-associated SNPs with cis-regulated e QTL effects. They regulated the expression of 9 e QTL genes, all these e QTL genes enrich in immune related pathways or Gene Ontology(GO) terms. Functional prediction analysis identified 4 SNPs(rs3129882, rs3135338, rs9268645 and rs3825932) with significant regulation effect(Regulome DB score). Of the 9 regulated genes, 6 genes(TAP2, HLA-DOB, HLA-DQB1, HLA-DQA1, HLA-DRB5 and CTSH) were differentially expressed in immune related cells. Of note, one SNP rs3825932 cis-regulated the expression of CTSH gene, which were in turn differentially expressed in PBMC, implying the causal effect of rs3825932 on the disease was through regulating CTSH expression. This hypothesis was also supported by subsequent functional prediction analysis, which showed that rs3825932 played an important role in chromatin structure formation and histone modification. In a word, the integrative functional evidence towards CTSH demonstrated that rs3825932 or other loci within CTSH participate in the development of T1 DM.Conclusions: By performing comprehensive integrative analysis of multi-omics genetic data, this study had exhaustively explored the possible functional mechanisms of T1 DM associated genetic variants. Our findings showed that some loci affect the development of T1 DM through their cis-regulation effect of relevant e QTL genes. As a proof-of-concept, an example of rs3825932 regulating CTSH was investigated in detail. Utilization of public data resources for integrative analyses may provide novel insights into the molecular genetic mechanisms underlying human disease.